U.S. patent application number 10/597092 was filed with the patent office on 2008-09-11 for method and kit for the production of particles labelled with rhenium-188.
This patent application is currently assigned to ROTOP PHARMAKA GMBH. Invention is credited to Antje Drews, Gerd Wunderlich.
Application Number | 20080219923 10/597092 |
Document ID | / |
Family ID | 34813101 |
Filed Date | 2008-09-11 |
United States Patent
Application |
20080219923 |
Kind Code |
A1 |
Wunderlich; Gerd ; et
al. |
September 11, 2008 |
Method and Kit for the Production of Particles Labelled with
Rhenium-188
Abstract
Rhenium-188 labeled particles are produced by suspending
particles of an organic polymer or a biopolymer in a solution
wherein the solution contains a water-soluble tin-II salt and a
Re-188 perrhenate salt with a radioactivity of 1,000 MBq to 60,000
MBq and wherein the solution has initially a pH value of pH 1 to pH
3; heating the solution of step a) to 80.degree. C. to 100.degree.
C.; and, after 45 minutes to 70 minutes of heating, increasing and
adjusting the pH value to a pH value of pH 5 to pH 8.5.
Inventors: |
Wunderlich; Gerd; (Dresden,
DE) ; Drews; Antje; (Dresden, DE) |
Correspondence
Address: |
GUDRUN E. HUCKETT DRAUDT
SCHUBERTSTR. 15A
WUPPERTAL
42289
DE
|
Assignee: |
ROTOP PHARMAKA GMBH
Radeberg
DE
|
Family ID: |
34813101 |
Appl. No.: |
10/597092 |
Filed: |
January 27, 2005 |
PCT Filed: |
January 27, 2005 |
PCT NO: |
PCT/DE05/00140 |
371 Date: |
August 28, 2006 |
Current U.S.
Class: |
424/1.69 ;
424/1.65 |
Current CPC
Class: |
A61P 35/00 20180101;
A61K 51/1251 20130101; A61P 35/04 20180101 |
Class at
Publication: |
424/1.69 ;
424/1.65 |
International
Class: |
A61K 51/08 20060101
A61K051/08; A61K 51/00 20060101 A61K051/00; A61P 35/00 20060101
A61P035/00 |
Foreign Application Data
Date |
Code |
Application Number |
Jan 29, 2004 |
DE |
10 2004 005 280.8 |
Claims
1-15. (canceled)
16. A method for producing rhenium-188 labeled particles, the
method comprising the steps of: a) suspending particles of an
organic polymer or a biopolymer in a solution wherein the solution
contains a water-soluble tin-II salt and a Re-188 perrhenate salt
with a radioactivity of 1,000 MBq to 60,000 MBq and wherein the
solution has initially a pH value of pH 1 to pH 3; b) heating the
solution of step a) to 80.degree. C. to 100.degree. C., c) after 45
minutes to 70 minutes of heating, increasing and adjusting the pH
value to a pH value of pH 5 to pH 8.5.
17. The method according to claim 16, wherein in step c) a solution
of citrate, acetate, or tartrate is used for increasing the pH
value.
18. The method according to claim 16, wherein in step c) a solution
of potassium sodium tartrate is used.
19. The method according to claim 16, wherein the solution of step
a) contains a complexing agent for stabilizing the tin-II salt,
wherein the complexing agent is selected from 2, 5-dihydroxy
benzoic acid acetic acid, citric acid, malonic acid, gluconic acid,
lactic acid, hydroxy isobutyric acid, ascorbic acid, tartaric acid,
succinic acid, salts of said acids, or glucoheptonate.
20. The method according to claim 16, wherein the solution of step
a) contains 2,5-dihydroxy benzoic acid as a complexing agent for
stabilizing the tin-II salt.
21. The method according to claim 16, wherein the particles have a
diameter of 10 .mu.m to 30 .mu.m.
22. The method according to claim 16, wherein initially the
water-soluble tin-II salt is present in the solution of step a) in
a concentration of 10 mmol/l to 50 mmol/l.
23. The method according to claim 16, wherein the particles are
comprised of human serum albumin.
24. A pharmaceutical kit for producing particles labeled with
Re-188, the kit comprising: a) a first container containing a
quantity of water soluble tin-II salt and a quantity of a
complexing agent for stabilizing the tin-II salt, the complexing
agent selected from 2,5-dihydroxy benzoic acid, acetic add, citric
acid, malonic acid, gluconic acid, lactic acid, hydroxy isobutyric
acid, ascorbic acid, tartaric acid, succinic acid, salts of said
acids, or glucoheptonate; b) a second container with particles made
from an organic polymer or a biopolymer; c) a third container
containing a quantity of a substance for increasing the pH value,
the substance selected from citrate, acetate, or tartrate, present
in solid form or in aqueous solution and generating in solution a
pH value of pH 6.5 to pH 8.5.
25. The pharmaceutical kit according to claim 24, wherein the
complexing agent for stabilizing the tin-II salt is 2,5-dihydroxy
benzoic acid.
26. The pharmaceutical kit according to claim 24, wherein the
substance for increasing the pH value is potassium sodium
tartrate.
27. The pharmaceutical kit according to claim 24, wherein the
particles have a diameter of 10 .mu.m to 30 .mu.m.
28. The pharmaceutical kit according to claim 24, wherein the kit
contains 0.02 mmol to 0.1 mmol of the tin-II salt per
administration to the patent.
29. The pharmaceutical kit according to claim 24, wherein the
particles are comprised of human serum albumin.
30. Rhenium-188 labeled particles produced by the method according
to claim 16.
31. Rhenium-188 labeled particles according to claim 30 as a
radiotherapeutic agent for treating tumors, carcinoma or their
metastases.
Description
[0001] The invention relates to a method for producing particles
labeled with radioactive isotope rhenium-188 (Re-188) and a kit for
performing the method. Such radioactively labeled particles can be
used in medicine, preferably in the field of oncology and nuclear
medicine, for radiotherapy of tumors or metastases of tumors.
[0002] The radiotherapy of tumors or their metastases with
radioactively labeled particles is known. In general, for this
purpose a catheter is inserted into the vessels leading to the
tumor. Through the catheter, the radioactively labeled particles
are subsequently supplied locally to the tumor tissue. The
radioactively labeled particles have a size that guarantees that
they get stuck when first passing the tumor-infiltrating capillary
blood system in the capillaries of the tumor. The method makes it
possible to reach very high radioactive doses in the targeted tumor
tissue while at the same time the surrounding tissue or other
organs of the patient are protected. Significantly higher radiation
doses in the tumor tissue have been achieved in comparison to e.g.
systemic intravenous application of radioactively labeled
antibodies, peptides and other low-molecular compounds.
[0003] In the last decade, primarily radionuclides Y-90, Re-188 and
Ho-166 have been used for labeling appropriate particles. The beta
ray emitter Re-188 with relatively minimal half-life of 17 hours is
especially suitable for a therapy with high radionuclide doses and
several applications to the same patient.
[0004] However, the current labeling methods for Re-188 and
particles are unsatisfactory.
[0005] Labeling methods that can be effectively performed in
related chemical elements, for example, technetium, are not
transferable onto labeling with Re-188 as a result of the different
chemical properties, in particular, the different redox
potentials.
[0006] Preferred as a carrier material in nuclear medicine for
radionuclide transport are human serum albumin microspheres of an
average particle size of 20 micrometers ([.sup.99mTc] HSA
microspheres B20, Rotop Pharmaka, Germany; Wunderlich G. et al.
Applied Radiation and Isotopes 52 (2000), pages 63-68). These
protein particles are degradable within the organism so that the
microspheres only temporarily clog the capillaries and can be
infused several times to the patient. When the labeling method
developed for technetium is used for labeling Re-188, labeling
yields of only less than 5% are achieved as a result of the
differences in the redox potential.
[0007] A disadvantage of the method described in Wunderlich et al.
is that after more than 90 min. reaction time only 70% to maximally
90% of Re-188 is bonded to the particles. In order to prevent that
unreacted Re-188 causes undesirable radiation exposure in the
organism of the patient, it is necessary to remove the excess
Re-188 by several washing steps. These washing steps require a
direct handling of radioactive liquids and therefore cause a high
radiation exposure of the personnel.
[0008] Publications by Wang S. J. et al. (Journal of Nuclear
Medicine 1998, 39 (10), pp. 1752-1757, Nuclear Medicine
Communications 1998, 19: pp. 427-433) also disclose methods for
labeling microspheres with Re-188. These microspheres are comprised
of plastic resin. Disadvantageously, in these methods the
microspheres after labeling with Re-188 must also be washed by
removing the supernatant and resuspending in saline solution. In
this method, for labeling 20 mg microspheres 200 mg tin salt and a
highly acidic pH value are required. The great amount of tin has
the disadvantage that the patient is additionally pharmacologically
exposed. Because of the highly acidic pH value, the method is not
suitable for protein particles because the protein would be
hydrolyzed by the employed highly acidic 0.2 N HCl.
[0009] Grillenberger K. G. et al. disclose Re-188 labeled hydroxy
apatite and sulfur colloid (Nuclear Medicine 1997, 36: pp. 71-75).
The yield obtained by this labeling method is however less than
80%.
[0010] The known methods for labeling particles with Re-188 are
time-consuming. The obtained labeling yields are greatly dependent
on the employed base material.
[0011] There is a need in nuclear medicine for a method that
simplifies labeling of particles with Re-188 for hospital
personnel. The handling of high radioactive doses of rhenium-188
should be as short as possible in order to keep the radiation
exposure of the personnel within an acceptable range. A method that
can eliminate washing steps would therefore be desirable.
[0012] It is an object of the invention to provide a simplified
method for labeling particles with rhenium-188 as well as a kit for
performing the method. In particular, the method and the kit should
reduce the radiation exposure of the personnel and the required
time for performing the method.
[0013] According to the invention, the object is solved by a method
for producing rhenium-188 (Re-188) labeled particles in which
method the particles are first suspended in an acidic solution and
heated and, after a certain amount of time of heating, the pH value
is increased.
[0014] The solution has in this connection a pH value of pH 1 to pH
3 and contains: [0015] a) a tin-II salt and [0016] b) a Re-188
perrhenate salt.
[0017] The preferred reaction volume in this step is 1 to 5 ml,
especially preferred 2 ml to 4 ml, especially advantageously 3 ml.
The known Re generators deliver a minimum eluate volume of 2 ml.
Advantageously, by means of the reaction volume in the method
according to the invention the entire eluate of a Re generator can
be utilized.
[0018] After 30 to 240 minutes, preferably 45 to 70 minutes, of
heating, the pH value is increased. In this connection, the pH
value is adjusted to be greater than pH 5, preferably between pH
6.5 to pH 8.5.
[0019] Surprisingly, the yield of labeling the particles with
Re-188 is increased to more than 95% by increasing the pH value at
the end of heating. By the thus obtained effective labeling of
particles with Re-188 a further processing of the end product is no
longer required. In particular, washing steps are no longer needed.
The suspension obtained by increasing the pH values can be directly
used for radiotherapy of the patient.
[0020] The total reaction time is shortened significantly in
comparison to the prior art. By eliminating washing steps, in
addition to saving time the radiation protection for the personnel
is significantly improved because fewer manipulations are required
in order to arrive at an injectable product.
[0021] By means of the method a specific radioactivity (labeling of
the particles) is reached that is significantly above the labeling
that has been described before by Wunderlich et al. (2001): 2,500
MBq/mg in comparison to 500 MBq/mg. The increase of the pH value is
realized by adding a buffer solution, preferably acetate, citrate,
or tartrate solution, especially preferred a potassium sodium
tartrate solution.
[0022] The buffer solution after having been added to the heated
solution preferably has a final concentration of 15 mmol/l to 50
mmol/l, particular preferred 25 mmol/l.
[0023] The tin-II salt is preferably a water soluble tin-II salt,
for example, SnCl.sub.2.times.2H.sub.2O or SnF.sub.2, which at the
beginning of the method is present in the solution in a
concentration of 10 mmol/l to 50 mmol/l, especially preferred 17
mmol/l.
[0024] By means of the method, the Re-188 initially present as
perrhenate (ReO.sub.4.sup.-) in the oxidation state +VII is reduced
by the reductive effect of the tin-II salt. In this way, the oxide
of Re-188 is precipitated in the oxidation state +4
(ReO.sub.2.times.H.sub.2O) together with the generated sparingly
soluble tin hydroxide on the microspheres. The resulting layer
generated by co-precipitation has a thickness of approximately 1
.mu.m.
[0025] With the method according to the invention, the amount of
the tin-II salt required for labeling can be reduced by a factor 10
in comparison to the prior art (Wang et al.). An amount of 10 mg to
12 mg of tin(II) salt per 10 mg microspheres has surprisingly been
found to be sufficient for labeling the microspheres.
[0026] Since tin-II salts are relatively instable in aqueous
solution when heated, a complexing agent for stabilizing the tin-II
salt is added to the solution. Such a complexing agent is
preferably an organic carboxylic acid, especially preferred
2,5-dihydroxy benzoic acid (gentisic acid). Further preferred
complexing agents are acetic acid, citric acidic, malonic acid,
gluconic acid, lactic acid, hydroxy isobutyric acid, ascorbic acid,
tartaric acid, succinic acid, the salts of the aforementioned acid,
or glucoheptonate. The complexing agent for stabilizing the tin-II
salt has in solution preferably a concentration of 50 mmol/l to 30
mmol/l, particularly preferred 20 mmol/l.
[0027] The use of gentisic acid is advantageous because gentisic
acid is a radical scavenger and therefore acts as a
radiation-protective agent in the preparation. Gentisic acid,
moreover, is already approved as an additive for
pharmaceuticals.
[0028] Heating of the solution is realized preferably to a
temperature below boiling point, in a range of 80.degree. C. to
100.degree. C.
[0029] The particles to be labeled are preferably spherical or
approximately round. Such particles, referred to as microspheres,
have advantageously a diameter that is small enough so that the
microspheres can be transported through normal blood vessels but
large enough that they get stuck in the capillaries. Preferably,
they have a diameter of 10 .mu.m to 100 .mu.m, especially preferred
15 .mu.m to 30 .mu.m.
[0030] The particles are preferably comprised of an organic polymer
or a biopolymer. In one embodiment of the invention, the particles
are comprised of a polymer that cannot be degraded in vivo,
preferably a weak cation exchange resin (e.g. Bio-Rex 70, BioRad,
Germany), polyacrylate, polymethylmethacrylate (PMMA, e.g. Heraeus
Kulzer, Germany), methacrylate copolymer (e.g. MacroPrep, BioRad,
Germany) or polyvinyl formaldehyde (e.g. Drivalon,
Nycomed-Amersham, Germany).
[0031] Particularly preferred particles are however microspheres of
a material that can be metabolized and degraded in the human
organism so that the particles will clog the capillaries after
application only temporarily. Advantageously, this enables several
applications of the particles. A preferred example of such
degradable particles are microspheres of human serum albumin
([.sup.99mTc] HSA microspheres B20, Rotop Pharmaka, Radeberg,
Germany). The [.sup.99mTc] HSA microspheres B20 are already
approved for use with labeling by technetium 99m.
[0032] Comparative examples with particles of different
biodegradable materials have shown that with microspheres of human
serum albumin in the method according to the invention surprisingly
a significantly higher Re-188 labeling yield and greater in vivo
stability can be achieved than with other materials that are also
degradable in vivo. For example, the labeling yield with particles
of macro-aggregated albumin (MAA, Nycomed-Amersham, Germany),
collagen particles (Angiostat, Regional Therapeutics, USA) and
polyacetate particles (PLA, Micromod, Germany) is significantly
lower.
[0033] The particles during labeling are preferably present in a
concentration of 2 to 3 million particles, preferably 2.5 million
particles, per milliliter, or 0.5 to 10 million particles per
milliliter.
[0034] The beta ray emitter rhenium-188 used for labeling is
practically available in unlimited quantities over several months
after purchasing a corresponding radionuclide generator (Oak Ridge
National Laboratory, TN, USA, or Schering AG, Germany) and is
suitable in particular for a therapy with high radionuclide doses
and several applications to the same patient. In such a generator,
the Re-188 is eluated in the form of perrhenate (oxidation state
VII of Re-188) by applying an 0.9% saline solution. The thus
obtained Re-188 generator eluate has preferably a radioactivity of
1,000 MBq to 60,000 MBq, preferably of 10,000 to 20,000 MBq.
[0035] The specific radioactivity (labeling of the particles)
obtained with the method according to the invention is preferably
1,500 to 3,000 MBq/mg. Advantageously, the specific radioactivity
can be adjusted in regard to the patient to the desired therapeutic
radiation dose by the employed amount of Re-188 generator
eluate.
[0036] Advantageously, the method according to the invention is
therefore suitable for labeling microspheres with radioactivities
that are within the therapeutic range. Because of this and because
of the aforementioned simplification of the method steps, the
development of a pharmaceutical kit is advantageously possible.
[0037] A further object of the invention is a pharmaceutical kit
for performing the method according to the invention. This kit for
producing rhenium-188 labeled microspheres comprises the following
components: [0038] a) a container with a quantity of water-soluble
tin-II salt and a complexing agent stabilizing the tin-II salt,
each in a powder form or as a solution, [0039] b) a second
container with microspheres of human serum albumin, as well as
[0040] c) a third container with a substance or solution for
increasing the pH value, in powder form or as a solution.
[0041] The substance for increasing the pH value is present in
solid form or aqueous solution and results in solution in a pH
value of pH 6.5 to pH 8.5.
[0042] Preferably, the components are distributed onto different
containers. The kit contains in this embodiment at least one of the
three containers per administration to the patient.
[0043] In an especially advantageous configuration of the
invention, acetate, citrate or tartrate, preferably potassium
sodium tartrate, is used for increasing the pH value. For each
administration to the patient, the kit contains preferably 0.1 mmol
to 0.2 mmol of a substance for increasing the pH value, especially
preferred 30 mg to 50 mg potassium sodium
tartrate.times.4H.sub.2O.
[0044] The tin-II salt is preferably a water-soluble tin-II salt,
for example, tin(II) chloride dihydrate or SnF.sub.2. For each
administration to the patient, the kit contains preferably 0.02
mmol to 0.1 mmol of the water-soluble tin-II salt, especially
preferred 5 mg to 20 mg tin(II)chloride dihydrate.
[0045] Because the tin-II salts in aqueous solution are relatively
instable when heated, the kit contains preferably as a further
component a complexing agent for stabilizing the tin-II salts. Such
a complexing agent is preferably an organic carboxylic acid or a
salt of an organic carboxylic acid. The complexing agent is
contained in the container (a) with the tin-II salt.
[0046] An especially preferred complexing agent for stabilizing
tin-II salt is 2,5-dihydroxy benzoic acid (gentisic acid). Further
preferred complexing agents are acetate, citrate, malonate,
gluconate, malate, lactate, hydroxy isobutyrate, pyrophosphate,
ascorbate, potassium sodium tartrate or glucoheptonate. For each
application to the patient the kit contains preferably 0.5 to 2
mol, in particular preferably 1 mol, of the complexing agent
stabilizing the tin-II salt per mol tin-II salt. This corresponds
to a quantity of 5 mg to 20 mg gentisic acid.
[0047] The kit contains as further components also the particles to
be labeled. These particles are preferably round or approximately
round. Such particles, microspheres, have advantageously a diameter
that is small enough that the microspheres can be transported
through regular blood vessels but large enough to get caught in
capillaries. Preferably, they have a diameter of 10 .mu.m to 50
.mu.m, especially preferred 10 .mu.m to 30 .mu.m.
[0048] The kit contains preferably 0.5 to 10 million, especially
preferred 1 to 5 million particles, advantageously 1 to 2 million
in an additional container (b).
[0049] The particles are comprised preferably of a material that is
metabolized and degraded in the human organism such that these
particles will clog the capillaries upon administration only
temporarily. Advantageously, in this way multiple applications of
the particles are possible. A preferred example of such degradable
particles are microspheres of human serum albumin ([.sup.99mTc] HSA
microspheres B20, Rotop Pharmaka, Radeberg, Germany). The
[.sup.99mTc] HSA microspheres B20 are already approved for use with
labeling by technetium 99m.
[0050] The particles are contained in the kit preferably in a
concentrated aqueous or alcoholic suspension. In order to increase
the dispersion of the particles, advantageously a non-ionic
detergent is added to this suspension. Preferably, non-ionic
detergents of the polyethylene type, for example, polyoxyethylene
sorbitan monooleate (Tween.RTM. 80), are used.
[0051] The non-ionic detergent is preferably contained in an amount
of 0.15 mg to 0.3 mg per 1 mg particle in the suspension.
[0052] For producing Re-188 labeled microspheres, the tin-II salt
and the complexing agent for stabilizing the tin-II salt are
dissolved in the first container in sterile water and added to the
second container containing the microspheres and the microspheres
are suspended in the solution. The generator eluate containing the
radioactive rhenium-188 is added to the suspension and the
suspension is heated to 80.degree. C. to 100.degree. C. After 45
minutes to 70 minutes of heating, the pH value is adjusted to pH 5
to pH 8.5 by mixing the suspension with the substance for
increasing the pH value that is contained in the third container.
The suspension is now cooled, preferably to body temperature, and
can be administered without washing steps directly to the
patient.
[0053] The invention also concerns the particles produced with the
method according to the invention and the kit according to the
invention and their use for radiotherapy of carcinoma or their
metastases.
[0054] A further component of the invention is a method for
radiotherapy of tumors, carcinoma or their metastases with these
particles. In the method, by means of the method as described above
particles labeled with Re-188 are produced. Into the local blood
vessel that leads to the carcinoma a catheter is inserted. Through
the catheter, a suspension of the radioactively labeled particles,
after adjusting the pH value to pH 5 to pH 8.5, is subsequently
supplied locally to the tumor tissue (without intermediate washing
of the particles). The radioactively labeled particles have a size
that ensures that upon the first passage of the tumor-infiltrating
capillary blood system they remain within the capillaries of the
tumor. Preferably, the particles have for this purpose a diameter
of 10 .mu.m to 50 .mu.m, particularly preferred 10 .mu.m to 30
.mu.m.
[0055] The method enables advantageously that very high
radioactivity doses are reached in the targeted tumor tissue while
at the same time the surrounding tissue and other 30 organs of the
patient are protected. Significantly higher radiation doses
(100-150 Gy) in the tumor tissue are achieved in comparison to, for
example, systemic intravenous administration of radioactively
labeled antibodies, peptides and other low-molecular compounds.
[0056] The use of microspheres of human serum albumin has the
advantage that the particles can be degraded in the body. The
microspheres close off the capillaries only temporarily when
administered. Multiple administrations are thus possible.
[0057] The administration of the particles is carried out
preferably arterially by means of infusion. For this purpose,
preferably 0.5 to 10 million, particularly preferred 1 to 5 million
particles, advantageously 1 to 2.5 million (corresponding to 1 to
20 mg, preferably 3 to 10 mg) are suspended in 20 ml to 100 ml,
preferably 50 ml, of an infusion solution (for example, sterile
isotonic saline solution) and infused.
[0058] The microspheres are degraded with a biological half-life in
the range of preferably greater than 200 hours, preferably eight
days to 15 days. The biological half-life of the microspheres is
thus in the range of the biological half-life of Re-188.
Advantageously, by immobilizing Re-188 on the microspheres, the
Re-188 is fixed at the application location (>90% remain
resident there over days).
[0059] The microspheres labeled with Re-188 in accordance with the
present invention are suitable advantageously in particular for the
therapy of liver carcinoma and liver metastases of other
carcinoma.
[0060] With the aid of the following examples the invention will be
explained in more detail:
EXAMPLE 1
[0061] Labeling of particles with Re-188 is explained with the aid
of labeling of human serum albumin (HSA) microspheres as
follows:
[0062] 9.3 mg 2,5-dihydroxy benzoic acid (gentisic acid) are
dissolved in 2 ml water for injection, subsequently 11.4 mg
SnCl.sub.2.times.H.sub.2O are added, and the solution is sterilely
filtered into a bottle containing human serum albumin (HSA)
microspheres (MS B20, Rotop Pharmaka, Radeberg, Germany). The
particles in the bottles are slurried and transferred into another
kit bottle MS B 20 and subsequently into a third kit bottle. In the
third bottle 1.5 million particles MS B 20 are then contained. To
this is added 1 ml sterilely filtered Re-188 perrhenate
(10,000-20,000 MBq) dissolved in 0.9% NaCl. The kit bottle with the
particles is then inserted into a heating block and the latter is
shaken for 55 minutes at 95.degree. C. Subsequently, 0.6 ml
sterilely filtered KNa tartrate solution (42 mg/ml) are added and
heating is switched off. After five minutes of additional shaking,
the preparation is ready to be injected.
[0063] The labeling yield (radiochemical purity) of the particles
labeled in this way is than 95%.
EXAMPLE 2
[0064] A preferred kit for labeling particles (in this case: human
serum albumin (HSA) microspheres) with Re-188 is comprised of three
flasks with the ingredients listed in Table 1.
TABLE-US-00001 TABLE 1 bottle component quantity/bottle process
consistency 1 2,5 dihydroxy benzoic acid 9.3 mg lyophilized solid
tin(II)chloride dihydrate 11.4 mg ultra high purity nitrogen 5.0 2
HSA microspheres A20 10 mg (1.2 .times. vacuum- solid (diameter
10-30 .mu.m) 10.sup.6 to 2 .times. 10.sup.6 concentrated particles)
Tween .RTM. 80 2.4 mg ultra high purity nitrogen 5.0 3
potassium-sodium tartrate 1 ml sterilized liquid solution (42
mg/ml) ** ultra high purity nitrogen is used as an inert gas
[0065] The kit is designed for the treatment of a patient.
EXAMPLE 3
[0066] With the kit according to Example 2 the particles (in this
case: human serum albumin (HSA) microspheres) are labeled according
to the following labeling procedure:
[0067] The components of the kit bottle 1 (2,5-dihydroxy benzoic
acid--gentisic acid) and tin(II) chloride dehydrate) are dissolved
in 2 ml sterile pyrogen-free water for injection purposes and added
in the kit bottle 2 to the HSA microspheres A20. After adding the
solution, for pressure compensation the same volume of nitrogen is
to be removed with a syringe from the bottles 1 and 2. By slight
shaking causing wetting of the rubber lyo stopper the HSA
microspheres are suspended.
[0068] [.sup.188Re] sodium perrhenate in sterile, isotonic
pyrogen-free sodium chloride solution (.sup.188Re generator eluate
(10,000-20,000 MBq), volume: 1 ml) is transferred into the bottle 2
which is arranged in a lead shielding. After adding the .sup.188Re
generator eluate for pressure compensation the same volume of
nitrogen is to be removed from the bottle 2.
[0069] For carrying out the reaction, the bottle 2 is shaken in a
heater/shaker for 55 minutes at 95.degree. C. The bottle 2 is
removed from the shaker and 0.6 ml of the bottle 3 (K/Na tartrate
solution) is transferred into bottle 2. After adding the solution,
for pressure compensation the same volume of nitrogen is to be
removed from bottle 2. By slight shaking with wetting of the rubber
lyo stopper the [.sup.188Re] HSA microspheres are suspended.
[0070] The preparation in the bottles is allowed to react for 5
more minutes at room temperature by using the shaker; the
preparation is then ready to be injected. The suspension of the
labeled [.sup.188Re] HSA microspheres B20, depending on the desired
concentration, can be diluted with sodium chloride solution for
injection. The [.sup.188Re] HSA microsphere suspension can be used
up to two hours after labeling.
EXAMPLE 4
[0071] The kit according to Example 2 is produced as follows:
[0072] For a batch of 150 bottles No. 1.395 g gentisic acid
(2.5-dihydroxy benzoic acid) and 1.710 g of tin(II) chloride
dihydrate are dissolved in 150 ml water for injection purposes. The
solution is distributed onto 150 bottles and lyophilized.
[0073] For a batch of 200 bottles No. 2, 2.0 g HSA microspheres A20
(Rotop Pharmaka GmbH, Germany) and 0.48 g Tween.RTM. 80 are
suspended in a solution of: [0074] 360 mm acetone [0075] 40 ml
sodium hydroxide solution (0.1 mol/l) [0076] 40 mm hydrochloric add
(0.1 mol/l [0077] 240 ml ethanol abs.
[0078] To the suspension a minimal amount of the dye bengal pink is
added. The suspension is concentrated in vacuum to 400 ml and
distributed onto the 200 bottles. Subsequently, acetone and ethanol
are removed by vacuum drying.
[0079] For a batch of 150 bottles No. 3, 6.3 g potassium sodium
tartrate are dissolved in 150 ml water for injection purposes. The
solution is distributed onto 150 bottles.
EXAMPLE 5
[0080] In accordance with the procedure of Example 1, particles of
different materials were labeled with Re-188:
S1 weak polyacrylate cation exchange resin (Bio-Rex 70, BioRad,
Germany), S2 polymethylmethacrylate (PMMA, Heraeus Kulzer, Germany)
S3 methacrylate copolymer (MarcoPrep Q, BioRad, Germany) S4
polyvinyl formaldehyde (Drivalon, Nycomed-Amersham, Germany) S5
macro-aggregated albumin (MM, Nycomed-Amersham, Germany) S6 human
serum albumin (HSA B20, ROTOP Pharmaka GmbH, Germany) S7 collagen
particles (Angiostat, Regional Therapeutics, USA), S8 polylactate
particles (PLA, Micromod, Germany).
[0081] 2-3 mg of the particles (corresponding to approximately 0.5
million particles) were used, respectively.
[0082] Before and after labeling, the distribution of the particle
size was determined according to ISO 13323-1 by means of
single-particle light scattering. After dilution in particle-free
water the particles were measured sequentially in the measuring
zone of the flow cuvette. The size distribution of the particles
was recalculated according to ISO 9276-2 into a surface area-based
distribution because this better characterizes the distribution of
Re-188 on the labeled particle surface.
[0083] In Table 2 the values of the cumulative distribution
(Q.sub.2) according to ISO 1998 is provided; the values represent
90% of the surface area-based total distribution.
[0084] The labeling yield was determined after labeling by
centrifugation of the particle suspensions and radioactivity
measurement of the supernatant and precipitate in an automated
gamma counter (Cobra II, Packard, USA).
TABLE-US-00002 TABLE 2 particle size particle before size after
labeling labeling labeling yield material [.mu.m] [.mu.m] [%] S1
Biorex 70 macro reticular acrylic 45-75 30-75 80-85 resin S2 PMMA
polymethylmethacrylate 4-25 4-25 70-85 S3 Macro Prep methacrylate
Copolymer 45-55 30-80 83-90 S4 Drivalon polyvinyl formaldehyde
50-150 5-150 60-70 S5 MAA human serum albumin 10-100 10-50 60-70 S6
HSA B20 human serum albumin 13-27 15-37 95* S7 Angiostat collagen
20-75 1-15 35-50 S8 PLA polylactate 10-45 3-45 50-60
[0085] After labeling with Re-188 the biodegradable HSA
microspheres B20 (under the microscope recognizable as spheres) had
a hardly changed distribution between 15 .mu.m and 37 .mu.m
(average value 21 .mu.m).
[0086] In contrast to this, the macro-aggregated HSA (MAA) after
labeling had a broad particle distribution. This is caused by MAA
particles not being present as round microspheres but having
irregularities similar to little sponges. MAA particles are not so
stable at high temperatures and, because of the greater surface
area, are also much faster attacked and degraded enzymatically in
vivo.
[0087] Drivalon (S4), Angiostat (S7) and PLA (S8) particles also do
not survive well the labeling process at the required high reaction
temperature i.e., there is increased fine material and the particle
distribution is broadened significantly. Despite of this, labeling
for all particle preparations in vitro is rather stable.
[0088] In order to test the in vitro stability, the labeled
particle samples were incubated with human plasma. After three
hours of incubation at 37.degree. C. or after 24 h and 48 h
incubation at room temperature, the adhesion of Re-188 on the
particles after centrifugation and radioactivity measurement was
determined in an automated gamma counter (Cobra II, Packard,
USA).
[0089] The results of in vitro stability of the labeled particles
are summarized in Table 3.
TABLE-US-00003 TABLE 3 particle-bonded radioactivity, average value
.+-. standard deviation (SD) [%] t = 3 t = 24 t = 48 material t = 0
h/37.degree. C. h/22.degree. C. h/22.degree. C. S1 Biorex 70 100
93.3 .+-. 2.3 92.3 .+-. 1.6 86.3 .+-. 3.5 S2 PMMA 100 95.2 .+-. 1.7
93.8 .+-. 2.4 91.3 .+-. 2.7 S3 Macro Prep 100 92.7 .+-. 3.4 83.5
.+-. 1.6 82.1 .+-. 2.8 S4 Drivalon 100 95.0 .+-. 2.5 84.4 .+-. 3.4
79.9 .+-. 3.5 S5 MAA 100 97.3 .+-. 2.0 92.1 .+-. 1.7 86.3 .+-. 3.1
S6 HSA B20 100 98.0 .+-. 1.8 92.2 .+-. 2.2 86.8 .+-. 2.4 S7
Angiostat 100 92.0 .+-. 3.4 85.2 .+-. 3.2 82.8 .+-. 1.6 S8 PLA 100
96.1 .+-. 2.7 80.4 .+-. 2.8 75.7 .+-. 1.9
[0090] The in vitro stability of all particle preparations can be
considered to be satisfactory because 75-90% of Re-188 after 48
hours is still particle-bonded (Table 3).
[0091] The biodistribution of the different particles was examined
in vivo after intravenous injection in Wistar rats, wherein the
lungs served as a model for a tumor that has a good blood
supply.
[0092] After injection of particles labeled with 20 MBq Re-188 the
biodistribution of the particles was examined over 48 hours under
gamma camera (Picker CX 250) with the aid of conventional
nuclear-medical imaging technology. At the end of the gamma camera
examinations, the animals were killed, select organs removed, and
their radioactivity determined in a gamma counter in comparison to
the entire animal and to the injected activity.
[0093] The in vivo biodistribution in the liver and the lungs (in %
of injected doses in the entire organ, respectively) of the labeled
particles was determined 48 h after injection into the tail vein of
8-week old Wistar rats (n=3 to 6) for each material.
[0094] In order to determine the in vivo stability of the different
particle preparations, the biological half-life in the lungs
(T.sub.b 1/2) was used as a gauge and delivered values between 45
hours up to more than 200 hours. A biological half-life of 200
hours corresponds in this connection to an effective half-life of
15.4 hours for Re-188. Since after five effective half-lives (i.e.,
77 hours) only approximately 3% of the initial radioactivity is
present in the body and can act therapeutically, the obtained
stability can be considered to be satisfactory.
[0095] The results of in vivo biodistribution and in vivo stability
are summarized in Table 4.
TABLE-US-00004 TABLE 4 liver liver (% injected (% injected material
dosage) dosage) T.sub.b 1/2 [b] S1 Biorex 70 3.9 91.4 >200 S2
PMMA 16.7 76.4 >200 S3 Macro Prep 0.9 85.1 >200 S4 Drivalon
56.5 19.6 125.3 S5 MAA 2.8 48.0 45.4 S6 HSA B20 0.8 92.9 >200 S7
Angiostat 49.6 14.5 129.7 S8 PLA 11.1 66.5 153.9
[0096] The bio distribution studies show very good in vivo
stability for the preparations S1 to S3 and S6, characterized by a
very slow drop of radioactivity in the lungs and a minimal
radioactivity uptake in the non-target tissues (for example, the
liver, except in the case of S2--Table 4). S2 has already in the
base material a relatively high proportion of fine particles that
leads to particle deposition in the liver where the particles
however stay for the duration of the experiment and remain
unchanged.
[0097] Small particles (<10 .mu.m, as, for example, sample S2)
are collected in reticulo-endothelial system (RES) in liver and
spleen. When fine material is generated in the labeling process, it
is found after intravenous (iv) injection in these organs (sample
S4, S7, and S8). These particles are therefore not suitable for
intra-arterial tumor therapy in humans even though the biological
half-life is relatively long (>120 h).
[0098] MAA macro-aggregates (S5) are not suitable for
intra-arterial tumor therapy in humans because of their relatively
minimal biological half-life (45.4 h).
[0099] In comparison to the fatal medical results reported by
Mantravadi 1981 (Mantravadi R V, Spigos D G, Tan W S, Felix E L,
Intraarterial yttrium-90 in the treatment of hepatic malignancies;
Radiology 1981; 142: 783-786) when employing a long-lived beta
emitter (Y-90) not satisfactorily bonded to the particles, the
application of particles labeled with short-lived emitter Re-188 is
significantly less dangerous. Re-188 released by the particles is
not accumulated in vitally important organs but in a short period
of time is excreted via the kidneys.
[0100] A further advantage of the use of Re-188 preparations is
that the available radionuclide generator can be employed at any
time for producing Re-188 preparations so that a request by a
physician can be responded to without a long waiting period and at
attractive costs.
[0101] The results of the comparative examples with different
particle materials can be summarized as follows:
[0102] With the method according to the invention different
particle materials can be labeled in high yield with Re-188 and
used in a promising way for endo-radiotherapeutic applications.
[0103] HSA microspheres B20 labeled with Re-188 are the most
attractive nuclear medical therapeutic agent for a local tumor
therapy after selective catheter placement in the supplying blood
vessels, in particular because of the biocompatibility of the
particles, their uniform size, and because of the high in vivo
stability of the product.
* * * * *